Project Summary/Abstract Coronary artery disease, as a result of atherosclerosis, is a major cause of debilitation and death, affecting millions of Americans. Despite decades of research, there are still large gaps in our understanding of the disease. For one, RNA expression signatures of genes, long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) that are altered in atherosclerosis have not been adequately evaluated in human tissues. New insight into these expression changes can be coupled with genetic studies and animal studies to provide a clearer framework to the causes and potential treatments of atherosclerosis. The new, deep resource of the GTEx (Genotype Tissue Expression) NIH Roadmap Project provides an opportunity to evaluate expression data from over 800 coronary artery segments that have a full range of atherosclerotic plaque types. As recently described, the most accurate way to interpret tissue-level expression data is to statistically correct for the cellular composition of each tissue to then uncover cell-level changes in expression. We hypothesize that cell- specific expression patterns of genes and their regulatory elements will implicate disease-causing pathways in subtypes of macrophages and smooth muscle cells in atherosclerosis. The project will elucidate gene, miRNA, and lncRNA expression alterations in atherosclerosis in GTEx coronary artery tissues after performing a computational cellular composition correction. First, the project will generate cell-specific expression data on primary cells necessary for robust deconvolution methods to uncover expression alterations. After correct of the components of each cell type, each individual RNA will be evaluated for over- or under-expression in atherosclerosis. Then important genes and lncRNAs will be localized using RNA in situ hybridization to specific subtypes of macrophages and smooth muscle cells in plaque microenvironments. miRNAs will be localized with a novel expression microdisseciton technique. Finally, the expression and localization data will be integrated with genetic (GWAS) data to explore causal regulatory pathways in atherosclerosis. Altogether, this project will redefine the expression alterations that occur in human atherosclerosis and will provide a great resource to the wider scientific community.